Sandbox 33

Hen Egg-White Lysozyme

=Background= Hen Egg-White Lysozyme is an enzyme that was first described by the Russian scientist P. Laschtchenko in 1909 and its structure was solved for in 1965 by David Phillips via X-ray crystallography (Garrett, & Grisham, 2005). It is a single polymer of 129 residues (14.3kDa in weight) that catalyzes the hydrolysis of the polysaccharide wall of bacterial cells, breaking the β (1-4) linkage between N-acetylmuramic acid (NAM) and N-acetylglucosamine between the D and E sugars. The D and E sugars are the fourth and fifth sugars, respectively, in reference to the six sugars (oligosaccharide), identified by the letters A-F, that function as the ligand and bind to lysozyme. In hens (Gallus gallus), lysozyme is heavily concentrated in the egg white, serving as an anti-biotic as well as a nutrient to the developing eggs. Lysozyme is not only found in hen egg-whites but has many homologs that occur in a wide variety of organisms including humans. The gene for lysozyme in hens is expressed in the oviduct and in macrophages, which directly reflects its purposes (i.e. nutrition and defense). The one gene is controlled at the level of transcription by different means for the different locations and resulting functions (Worthington, 2010).



=Structure=

The Hydrophobic Effect
There are many levels of organization that contribute to protein stability, the strongest of which being the hydrophobic effect. The reason the hydrophobic effect is so powerful in determination of the overall protein structure is because proteins exist and function in solvents. Since the body is mostly water, regions that are polar like water is, hydrophilic regions, will congregate near it, while regions that are not polar will repel from it, will try to distance themselves from water by being isolated towards the inside or orienting towards other hydrophobic groups so that they might stabilize (Red=Oxygen, Green=Ligand, Purple=Polar, Grey=Hydrophobic). With few exceptions, water stays out of the Hydrophobic regions of the enzyme.This is all a result of entropy because it takes more energy for water to surround and stabilize hydrophobic regions (Garrett, & Grisham, 2005). However, if it does mosey over it will for what is called putative water bridges. They are stabilized by the active site and usually required in the mechanism.

van der Waals Forces
The few hydrophobic residues that do exist on the surface are in the active site, the region in contact with the ligand, and a stabilized by van der Waals forces. Van der Waals forces also exist within the enzyme and stabilize the enzyme of its own need.

Primary, Secondary, and Tertiary Structure
The primary structure 129 residues of 1hew result in a secondary structure of 5 alpha helices and 5 beta sheets. They are mostly stabilized by Hydrogen bonding between secondary elements, nitrogen, oxygen, hydrogen’s on polar particles and water. We can see in Jmol that even if the secondary elements aren't there, the Hydrogen bonding provides the scaffolding to support the enzyme. While not as common as Hydrogen bonding, an ionic bridge can help stabilize hydrophilic regions trapped in the hydrophobic interior. Their opposite charges satisfy one another. But not only is the secondary structure of 1hew stabilized by Hydrogen bonding, but the ligand is held secure and safe by it as well. The tertiary structure of Lysozyme is made up of several binding motifs. An antiparallel β-sheet occurs between a pseudo β -sheet from bases Lys1-Phe3 and Phe38-Thr40. A helix loop helix occurs from Cys80-Leu84, with a loop occurring from Ser85-Ile88 followed by another helix Thr89-Val99. Also, a β-ladder exists from the antiparallel arrangement of 3 β-sheets from Gln41-Thr47, Gly49-Ile55, and Leu56-Arg61. Each other these structures help reduce the strain on the enzyme.

Disulfide Linkages
Next, we will observe the disulfide bonds, shown as yellow bars with the bases the join noted. They occur between the following Cystine residues: 6 and 127, 30 and 115, 76 and 94, 64 and 80. They give strong support in the enzyme between pieces of the same chain.

=Activity=



Functional Preferences
The function of lysozyme is optimal under physiological conditions of a pH 6-9 with maximal function observed at pH= 6.2 and temperatures around 37C. Furthermore, while lysozyme can lyse short saccharides, it is more efficient when cutting 3 repeating NAG-NAM units (Worthington, 2010).

Mechanism
The <scene name='Sandbox_33/Active_site/3'>active site of Lysozyme has a few key components that are integral parts of its catalytic ability. Glu35 acts as an acid, donating an H+ to the O in the glycosidic bond. Asp52 will covalently catalyze the reaction by binding its carboxyl group to the unstable positive ion. Water then enters the system and a hydroxyl group will add to the sugar of the NAM. Both Glu35 and Asp52 will return to their natural states and will continue as catalysts.

=References=